The role of p16INK4a as a marker of HR-HPV and in the diagnosis of CIN has been well established, but its predictive value in the clearance of the virus after CIN treatment and its use as a prognostic marker of cervical cancer has not been studied. A series of 302 archival samples, including 150 squamous cell carcinomas (SCCs) and 152 CIN lesions, were subjected to immunohistochemical staining for p16INK4a and HPV testing using PCR with three primer sets (MY09/11, GP5+/GP6+, SPF). Follow-up data were available of 88 SCC patients, and 67 of the CIN lesions had been followed-up with serial PCR after conization. HR-HPV types were closely associated with CIN (OR 19.12; 95% CI 2.31-157.81) and SCC (OR 27.25; 95% CI 3.28-226.09). There was a significant linear relationship between the lesion grade and intensity of p16INK4a staining (p = 0.0001). The expression of p16INK4a was also closely related to HR-HPV (p = 0.0001). p16INK4a staining was a 100% specific indicator of CIN, with 100% PPV, and showed 83.5% sensitivity and 80.1% PPV in detecting HR-HPV. However, p16INK4a staining did not predict clearance/persistence of HR-HPV after treatment of CIN. Similarly, despite a slightly more favorable survival in women with strong/intense p16INK4a staining in univariate analysis, p16INK4a expression was not an independent prognostic predictor in multivariate survival (Cox) analysis. After adjustment for p16INK4a staining, HR-HPV, histological grade, International Federation of Gynecology and Obstetrics (FIGO) stage, and age, only the last two were significant prognostic predictors (p = 0.0001 and p = 0.003, respectively). The present data confirm the role of p16INK4a as a highly specific marker of CIN and HR-HPV type, but expression of this protein does not seem to be of any prognostic value in cervical cancer or in predicting the clearance of HR-HPV after treatment of CIN. We speculate that different subgroups of cervical cancer are characterized by aberrant p16INK4a/cyclin D/Rb pathways that are due to different mechanisms that can be mutually exclusive.